Cloning and expression of BDNF protein in Escherichia Coli

Background and Aim: Brain derived neurotrophic factor (BDNF) located on chromosome ۱۱p۱۴.۱, is a member of the nerve growth factor family which involved in the growth, differentiation, and survival of specific types of developing neurons both in the central and the peripheral nervous system. It is also involved in regulating synaptic plasticity in the central nervous system. Expression of a similar gene in human is reduced in both Alzheimer's and Huntington disease patients. This has been found to be related with many biological functions, including NMDA receptor activity, synapse stability, dopaminergic, cholinergic, serotonergic, and GABAergic signaling, synaptogenesis, and dendritogenesis. Due to the importance of BDNF in pharmaceutical industries as well as its application on neurological disorders treatment, producing BDNF is crucial and is considered as an important step in development of pharmaceutical industries.Methods: BDNF gene was optimized based on E. coli strain BL۲۱ expression system and synthetized in pUC ۵۷ vector, Then it was sub-cloned into pET۲۱a between NdeI and XhoI restriction sites. The constructed recombinant plasmid was transformed in to E.coli strain BL۲۱ and expressed under different condicions. Then BDNF protein was purified by affinity chromatography by nickel-agarose column. The protein purity was assayed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). Finally, its activity will be evaluated.Results: The highest protein expression level was determined with ۰.۵ mM of Isopropyl β-D-۱-thiogalactopyranoside (IPTG), at ۱۸°C in ۵ h of incubation. The expressed protein then was purified by Ni-agarose affinity chromatography and was illustrated by SDS-PAGE.Conclusion: In this study we have cloned and then expressed BDNF protein in E. Coli strain BL۲۱ and the purified protein then was achieved by Ni-agarose affinity chromatography.

کلیدواژه ها:

Brain-Derived Neurotrophic Factor ، Drug industry ، Escherichia coli